The therapeutic GVAX vaccine boosts the body’s immune system T-cells to fight pancreatic cancer. To identify clinical correlates of polyfunctional response of T-cells that produce 2+ cytokines per single cell to GVAX vaccine, we employed 32-plex single-cell cytokine profiling platform to evaluate T-cell polyfunctionality in patients with pancreatic cancer who had GVAX vaccination.
CD4+ T-cells were isolated with anti-CD4 microbeads from pre- and post-vaccination PBMCs and stimulated with anti-CD3/CD28 at 37°C, 5% CO2 for 24 hours. After stimulation, cells were loaded into a single-cell IsoCode chip containing ~12000 microchambers, each pre-patterned with a complete copy of a 32-plex antibody array. Protein secretions from ~1000 individual CD4+ T-cells were analyzed after 16-hour-on-chip incubation. Polyfunctional profile was assessed by the 5 functional groups: effector (Granzyme B, IFN-γ, MIP-1α, Perforin, TNF-α, TNF-β), stimulatory (GM-CSF, IL-2, IL-5, IL-7, IL-8, IL-9, IL-12, IL-15, IL-21), regulatory (IL-4, IL-10, IL-13, IL-22, TGF-β1, sCD40L, sCD137), inflammatory (IL-1b, IL-6, IL-17A, IL-17F, MCP-1, MCP-4), and chemoattractive (CCL-11, IP-10, MIP-1β, RANTES).
The single-cell analysis demonstrates a marked upregulation of polyfunctional CD4+ T-cells across 5 patients after GVAX vaccination compared to pre-vaccinated CD4+ T-cells. The enhanced polyfunctional strength index (PSI) of CD4+ T-cells by GVAX was predominated by antitumor-associated effector proteins including Granzyme B, IFN-Υ, MIP-1α, Perforin and TNF-α, mixed with small amounts of MIP-1β, sCD137 secretions. Most importantly, post- versus pre-vaccination fold-change of PSI was significantly associated with patient overall survival (P = 0.001), indicating a potential of PSI in predicting GVAX vaccine efficacy and outcomes of patients with pancreatic cancer.
Mackay S, Flynn B, Paczkowski P, Jaffee E, Zheng L, and Zhou J. Single-Cell Polyfunctionality of CD4+ T Cells Shows Promise as a Predictor of Overall Survival of Pancreatic Cancer Patients Treated with GVAX Vaccine. To be presented at FOCiS Annual Meeting 2018.
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